Microorganisms in soil assimilate, transform, and mineralize soil C to support growth. There are an estimated 2.6 × 1029microbial cells containing 26 Pg C in soils worldwide. Consequently, quantifying microbial growth in soil is critical for determining the degree to which microorganisms contribute to the global C cycle. Measuring taxonspecific microbial growth enables understanding of the contribution of microbial taxa to elemental transformations across ecosystems and their susceptibility to environmental perturbations. These measurements in soil have largely been lacking due to inadequate methods. Quantitative stable isotope probing (qSIP) with H218O is used to measure taxon‐specific growth of microbial taxa in soil, an improvement compared with traditional stable isotope probing (SIP). In qSIP, DNA extracted from both a labeled (18O‐enriched water) and an unlabeled treatment is separated into numerous density fractions by isopycnic centrifugation, and target genes are quantified and sequenced in each fraction. The taxon‐specific DNA density shift and ultimately the isotopic composition (18O enrichment) is calculated for each taxon. Here we discuss methods and illustrate the procedure for quantifying microbial taxon‐specific growth in soil with qSIP using heavy isotope enriched H218O.
Water is the medium of life, is involved in biochemical reactions, and is exchanged among internal pools and with the water in the external environment of organisms. Understanding these processes can be improved by isotopically labeling the metabolic water that is produced inside the cells of organisms during aerobic respiration.
Here we describe a new method for isotopically labeling cellular water by incubating microbes and plant tissues in air enriched in18O2. As oxygen gas is reduced during respiration, H218O is produced. The rate of H218O production and the synthesis of biomolecules that incorporate18O from H218O can be quantified using cavity ringdown spectrometry and isotope ratio mass spectrometry.
For
Any organism with aerobic respiration is amenable to labeling with18O2, and the method described here enables a new approach to investigate questions regarding plant and microbial physiology. In plants,18O introduced as metabolic water could be tracked as it moves between living cells and exchanges with external water. For probing soil microbial physiology, the method described here has the advantage over the application of exogenous H218O of not increasing the soil moisture, a disturbance that can affect microbial metabolism.
- PAR ID:
- 10395848
- Publisher / Repository:
- Wiley Blackwell (John Wiley & Sons)
- Date Published:
- Journal Name:
- Rapid Communications in Mass Spectrometry
- Volume:
- 37
- Issue:
- 6
- ISSN:
- 0951-4198
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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